ENZYME THERMISTOR ANALYSIS OF KINETICS AND STABILITY OF IMMOBILIZED INVERTASE

The kinetic behaviour of immobilized invertase was investigated using calorimetric measurements in an enzyme thermistor device. The hydrolys is of sucrose is an exothermic process with inhibition-like kinetics a bove 300 mmol/l; glucose gave no measurable heat of reaction and fruct ose molecules are condensed in an endothermic process. The concentrati on dependence of the temperature signal of fructose condensation is li near up to 2 mol/l at an optimal pH of 3.3, and the absolute value of the heat of reaction is only 25 % compared with that of the hydrolytic reaction. A higher K(m)-value and a lower V(max) are therefore assume d. The reaction heat of hydrolysis of sucrose decreases with increasin g product concentration. A shift of the equilibrium by increasing conc entrations of both products is accompanied by a competitive condensati on reaction in the case of fructose. The stability of invertase in aqu eous ethanol solutions was also investigated. It depends on the ethano l concentration and on the time of contact with the solvent. At 30 % e thanol no inactivation within 20 min. occurs, whereas at 60 % ethanol a linear dependence of the inactivation on the time of contact was fou nd. Above 80 % ethanol less than 10 % enzymatic activity remained afte r a five-minute treatment. This irreversible inactivation has to be co nsidered if invertase is applied in water miscible organic solvents.